This proposal is concerned with various aspects of mechanisms of regulation of lipoprotein lipase (LPL) in cell culture systems and whole organism. The following hypotheses will be tested: (1) Triglyceride rich lipoproteins regulate the synthesis and secretion of LPL from parenchymal cells. Hormones (insulin, glucagon, glucocorticoid) also modulate synthetic rates of LPL. (2) Post-translational modifications of secreted enzyme protein via phosphorylation-dephosphorylation reactions, and/or proteolytic cleavage lead to the formation of catalytically active enzyme which binds to the endothelium. (3) Hydrolysis of VLDL and portomicron triglyceride is an ordered process favoring hydrolysis of the most unsaturated triglyceride species and leading to a continuous enrichment of saturated species. (4) Unsaturation of the endothelial plasma membrane affects the specific activity of LPL. Testing of the previous hypotheses will be made possible because of availability of highly purified LPL, radioimmunoprecipitation technique and sensitive solid phase radioimmunoassay for LPL which detects both catalytically active and inactive species. Changes in enzyme specific activity, synthesis rates and secretion rates, in response to substrate concentrations, metabolic intermediates and hormones will be determined in preadipocytes and granulosa cells in culture. The specific activity and molecular properties of the intracellular, secreted and endothelial bound LPL protein will be determined. The presence of covalently bound phosphate in LPL will be confirmed, and correlation between dephosphorylation and activation established. The intermolecular specificity of LPL with respect to triglyceride unsaturation will be studied in vivo by following changes in triglyceride species composition in plasma, following injection of a triglyceride load in hepatectomized roosters. The role of the fatty acid composition of the endothelial cell plasma membrane or liposomes on the binding and catalytic efficiency of LPL will be determined. LPL will be localized in various tissues by electron-microscopy following treatment with high affinity (125I) anti-LPL immunoglobulin or with ferritin-labeled second antibodies.